The availability of blood products requires, for their use for therapeutic or nontherapeutic purposes, purification techniques which make it possible to obtain products of high purity, and preferably free of contaminants, in particular of viral contaminants.
In blood products, the viral contaminants may be enveloped viruses (HIV, hepatitis B, C, D, E and G viruses, and the like) or nonenveloped viruses (hepatitis A virus, parvovirus, and the like).
For many years, various international or national bodies have introduced increasingly strict standards for the preparation of blood products so as to prevent their application for therapeutic or nontherapeutic purposes when they contain viral contaminants (Council Directives 65/65 EEC, 75/319/EEC & 89/381 EEC).
At the European level, the CPMP standards (CPMP/BWT 268/95 and 269/95) require the use of certain treatments against enveloped or nonenveloped viruses.
It is in particular mentioned in these documents that an inactivation step using heat (dry or steam) or using pasteurization in the preparation of clotting factors is effective against the hepatitis A virus, but would not be very effective against other nonenveloped viruses, in particular parvoviruses.
On the other hand, a treatment comprising a chemical inactivation step (by addition of solvents-detergents) is effective for enveloped viruses but ineffective for treating nonenveloped viruses.
It is also known to use certain chemical agents such as beta-propiolactone, which is effective in the treatment of nonenveloped viruses but has the disadvantage of modifying the treated proteins.
It is also known that certain long treatments using pH modification, below pH 4, or the addition of proteases allows activation of some nonenveloped viruses such as parvoviruses. However, these treatments also modify the conformation and structure of the treated proteins.
Consequently, it is known that, to date, the majority of the physicochemical treatment steps capable of being used to obtain viral inactivation of blood products are either highly toxic, or unacceptably affect the conformation of the treated proteins, or are ineffective for treating nonenveloped viruses, in particular parvoviruses.
Parvoviruses are small nonenveloped DNA viruses which infect numerous animal species, including man (Handbook of Parvoviruses, Vol. 1, pp. 1-30, Disinfection, Sterilization and Preservation, Fourth Edition, Seymour S. Block, Ed. Lea & Febiger, Philadelphia-London). They are endemic in nature and cause a wide variety of diseases whose appearance largely depends on the state of development of the host.
Among these, parvovirus B19 is the only known member of the Parvoviridae family which is pathogenic for man. Likewise, murin parvovirus Hi can also infect man.
Parvovirus B19 infection in a healthy man may be asymptomatic or may induce benign diseases (example: fifth disease in children).
On the other hand, in immunodeficient patients or patients suffering from blood disorders, it may lead to chronic anaemias and to transient aplasias which may be associated with haemolytic anaemias.
Passing through the placenta, it may cause intrauterine death. It exhibits a remarkable tropism for the erythroid lines of human haematopoietic progenitor cells.
A recent epidemiological survey has shown that 50 to 60% of the adult French population and 36% of 1- to 15-year-old children have a positive parvovirus serology.
The process of viral DNA has been demonstrated by genetic amplification (PCR) in a number of batches of purified factor VIII concentrates, regardless of the methods of viral inactivation used.
This has been confirmed by the B19.sup.+ serology detection, without clinical sign, in 85% of haemophilic children who have received, since birth, only highly purified factor VIII concentrate (FVIII THPSD) used in France since 1988, free of any contamination with enveloped viruses (HIV, HBV, HCV) (Y. Lauriau et al., 1er congres de la Societe Francaise de Transfusion [1st conference of the French Transfusion Company] (1994)).
This observation indeed demonstrates that, without new methods of viral inactivation, targeted at the selective elimination of parvoviruses, in particular of parvovirus B19, from blood products, the probability of contamination is high.
Parvoviruses are extremely resistant, even at high temperature. Their haemagglutination properties and their infectivity are not affected by chemical treatments, such as chloroform or various acids, and most resist enzymatic digestions using RNase, DNase, papain or trypsin.